Invasive aspergillosis is one of the most feared diseases in the immunocompromised host, and its incidence is steadily increasing. Despite new therapies, the mortality of this disease is greater than 50%. A critical step in the pathogenesis o invasive aspergillosis is the interaction of Aspergillus fumigatus with pulmonary epithelial cells. After being inhaled, A. fumigatus conidia adhere to and are endocytosed by the epithelial cells that line the alveoli. In susceptible hosts, these conidia then germinate to form hyphae, which exit the pulmonary epithelial cells and invade the deeper tissues. During the previous project period, we identified key transcriptional regulators of A. fumigatus that govern both its interactions with host cells in vitro and virulence during invasive disease in immunosuppressed mice. Using transcriptional profiling, we uncovered potential target genes of these transcriptional regulators. Our preliminary data indicate that these target genes encode newly identified virulence factors that are important for the pathogenic interactions of A. fumigatus with pulmonary epithelial cells and the innate immune system. The experiments described in this proposal will identify and characterize two types of newly discovered A. fumigatus surface molecules and their host cell receptors. The first is A. fumigatus galactosaminogalactan (GAG), which is a novel pathogen-associated molecular pattern (PAMP) that likely plays important roles in both adherence to host constituents and modulation of the inflammatory response. The second set of fungal cell surface molecules consists of three A. fumigatus proteins that likely function as fungal ligands which mediate the pathogenic interactions of this organism with pulmonary epithelial cells. We will investigate both types of surface molecules by: (1) determining the roles of GAG and the candidate fungal ligands in host cell interactions in vitro; (2) elucidating the mechanisms by which they influence virulence in mouse models of invasive pulmonary aspergillosis; and (3) identifying their cognate host cell receptors. Furthermore, we will determine if and how GAG influences the function and surface expression of the fungal ligands, and whether the fungal ligands in turn influence the surface exposure of GAG and -glucan. Utilizing this body of research, our ultimate goal is to develop new therapeutic strategies to block A. fumigatus adherence and invasion. It is imperative to improve therapy for invasive aspergillosis because of the expanding population of immunocompromised patients who at high risk for this serious and life-threatening disease.